DESCRIPTION (Applicant's Abstract): Airway epithelial cells protect the lung against a battery of inhaled insults, but they may also contribute to physiologic impairment in disease. Despite this pivotal role, certain biologic properties of these cells remain poorly understood. Our goal is to elucidate progenitor-progeny relationships and identify airway epithelial stem cells. While many different cells contribute to cell renewal, we hypothesize that bonafide stem cells occupy specific epithelial niches. Testing this hypothesis is key for understanding phenotypic changes in airways disease, determining which cells are most at risk for transformation, and identifying targets for gene therapy. We will use both animals and cultured human airway epithelial cells as models and propose the following specific aims: 1. Test the hypothesis that mouse airway epithelial stem cells in vivo are confined to distinct morphological compartments. We will isolate basal, columnar and Clara cells from novel transgenic mice and will test their growth capacity and differentiation potential. 2. Test the hypothesis that human airway epithelial stem cells in vitro are confined to distinct morphological compartments. We will systematically dissect well differentiated human large and small airway epithelial cultures into constituent cell types and will test their growth capacity and differentiation potential. 3. Test the hypothesis that we can utilize differentiation competent human airway epithelial cell lines to model cell lineages and regulation of cell type specific differentiation. We will use novel immortalization strategies including interfering with the Rb tumor suppressor pathway and telomerization to create new airway epithelial cell lines capable of airway cell type-specific differentiation. The studies proposed in Aims 1 and 2 will establish the relationship between classic morphological cell type, expression of specific markers, proliferative capacity and differentiation potential. By rigorously testing the stem cell properties of specific airway epithelial cells, we will develop a model for cell lineages. Aim 3 will provide novel basic information regarding regulation of airway epithelial cell proliferation, differentiation, and death and will generate new research tools useful to address many basic issues in airway epithelial cell biology.